Understanding the function of commonly deregulated signaling pathways in human tumors is critical to elucidating the biological basis of cancer. The Erk MARK pathway is activated in cutaneous squamous cell carcinomas (SCCs) and can combine with G1 escape in experimental models to transform normal human skin into lethal SCC. Overactive Erk MARK signaling is well-recognized to produce uncontrolled proliferation and suppression of differentiation; however, the effectors that control these processes are largely unknown. Long noncoding RNAs (ncRNAs) represent newly discovered regulators of gene expression and fate. We have identified 9 novel ncRNAs within the epidermal differentiation complex (EDC) that are Erk MAPK- regulated. These ncRNAs, termed NEDCRs (noncoding EDC RNAs), display striking expression differences between normal and SCC tissue. Initial functional studies of the first member, NEDCR1, revealed potent effects on differentiation and neoplasia. These data show that NEDCR1 is a major mediator of epidermal differentiation and suggest a previously unreported role for long ncRNAs in this process. The broad goal of this Ruth L. Kirschstein NRSA Postdoctoral Fellowship (F32) application is to identify and characterize NEDCRs that are functionally important Erk MAPK effectors. These efforts will advance our understanding of the mechanisms that regulate epidermal homeostasis and neoplasia. In Aim I, we will test whether Erk MAPK-mediated inhibition of epidermal differentiation in neoplasia requires suppression of NEDCR1. shRNA and expression retrovectors for NEDCR1 will be developed and introduced into human epidermal tissue containing genetic elements known to transform skin into SCC. Well-defined and validated measures will be used to assess the effects of sustaining and depleting NEDCR1 in epidermal tumorigenesis. In Aim II, we will functionally assess the effects of NEDCR2-9 on epidermal homeostasis and neoplasia using the approach that enabled us to identify NEDCR1 as an important regulator of epidermal differentiation. siRNA oligonucleotides, along with shRNA and expression retrovectors, will be developed for NEDCR2-9 and introduced into cultured human skin. NEDCRs demonstrating an effect on epidermal growth and differentiation will then be studied in experimental models of epidermal neoplasia. PUBLIC HEALTH RELEVANCE: This proposal seeks to understand mechanisms controlling epidermal homeostasis and neoplasia, which are relevant to diseases exerting negative impacts on a large proportion of the U.S. population, including skin cancers and chronic wounds. The current efforts are designed to yield insight and targets to aid in the development of future therapies for these disorders.